This research program will investigate the cellular and molecular mechanisms of segmental pattern formation in annelids. Much of what is known about pattern formation in metazoans comes from studies investigating Hox genes whose products establish regional identity in bilaterians. However very little is known about the upstream regulation of Hox genes in systems outside of the highly specialized dipteran flies. Given the importance of Hox gene function in animals, the identification and characterization of the upstream regulators of Hox gene expression is central to our understanding of animal development. Our primary aim in the proposal is to characterize the developmental regulators of Hox gene activity in two annelid classes, leeches and basal polychaetes. This comparative approach will allow us to distinguish general developmental mechanisms of cell determination from phylum-specific ones and it will provide molecular insights into how morphological diversity is generated in annelids. The first goal of the proposal is to characterize the spatiotemporal distribution of segmental precursor cells in a basal annelid Capitella capitata. This would represent the first molecular and cellular characterization of such cells in a polychaete. The next two objectives are to isolate and characterize the regulatory gene)products that lie upstream of Hox gene regulation in polychaete and leech. To meet these objectives, we are ]adopting a candidate gene approach in combination with the construction and screening of cDNA subtraction libraries in both polychaetes and leeches. The characterization of segmentation genes in both annelid classes will provide insights into the mechanisms of pattern formation operating in animals.